Abstract
Introduction:
Data from clinical allogeneic bone marrow transplant (allo-BMT) and pre-clinical murine models of allo-BMT have shown that donor plasmacytoid dendritic cells (pDC) have important roles in regulating graft-versus-host disease (GvHD) and graft-versus-leukemia (GvL) activities of donor T cells. Using murine models of allo-BMT we have previously shown that 1) donor pDCs induce Th1 polarization of donor T cells and augment the GvL activity of T cells; and 2) the addition of pDC to grafts composed of purified T cells and HSC limited the subsequent development of GvHD. VIP is an immunosuppressive neuropeptide that regulates adaptive immune responses. We reasoned that VIP signaling may regulate activation of allo-specific T-cells, and the VIP pathway may be target for regulating GvHD and GvL in allo-BMT.
Methods:
To explore the mechanisms by which pDC and VIP signaling regulate T cell activation we used: 1) transgenic mice expressing GFP under the control of the VIP promoter to measure VIP expression in vivo in allo-BMT recipients; 2) one-way mixed lymphocyte reaction (MLR) to measure the proliferative response of transgenic luciferase positive T cells in response to allo-antigens via bioluminescence imaging (BLI); and 3) a model system of indirect presentation of allo-peptides derived from a H2-Ab MHC class II molecule by pDC to transgenic T cells expressing the TEa TCR. The effect of blocking vasoactive intestinal polypeptide (VIP) signaling during activation of allo-reactive T cells was assessed by using VIP-KO cells and by the addition of VIP peptide or a peptide antagonist of VIP (VIPhyb) to the one-way MLR. T cell proliferation and activation was measured by flow cytometry.
Results:
Analysis of VIP expression in donor pDC in murine models of allo-BMT showed >100-fold induction of VIP promoter activity in donor pDC and donor T cells during the first two weeks post-transplant, indicating that VIP expression in donor pDC may regulate T cell activation.Addition of endogenous native VIP suppressed T cell proliferation in one-way MLR but was reversed by addition of a 10x concentration of the VIP antagonist peptide (Figure 1B). Furthermore, adding 3 uM of the VIP antagonist to the MLR cultured significantly enhanced T cell proliferation. TEa peptide-primed T cells cultures with peptide-primed pDC from VIP knock-out mice had increased proliferation and expressed more of the activation markers CD69 and CD71 compared with T cells cultured with VIP-WT pDCs. Comparing pDC purified from marrow versus spleen, we found significantly more proliferation in T cells cultured with bone marrow VIP-KO pDCs than splenic VIP-KO pDCs, indicating that the less mature marrow pDC have greater antigen presenting ability than the more mature splenic pDC.
Conclusion:
These data suggest that 1) VIP is produced by donor pDC early after allo-BMT; 2) VIP inhibits T cell allo-proliferation in one-way MLR's; and 3) blocking VIP signaling using donor cells that cannot produce VIP or through the use of pharmacological inhibitors of VIP can augment activation and proliferation of T cells in response to indirect antigen presentation.The present findings support studies of VIP antagonist in allo-BMT to augment the GvL activity of T cells through indirect antigen presentation. Future studies include using the BLI analysis of MLR to determine the effect of novel drugs on T cell proliferation.
**Jingxia Li and Reema Panjwani were equal contributors to this abstract.
No relevant conflicts of interest to declare.
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